A porphyrin biosynthetic pathway serves for the synthesis of chlorophyll and heme which play a vital role in plant metabolism, and it takes place in the chloroplast. In this pathway, protoporphyrinogen IX oxidase (hereinafter, referred to as PPO; EC:1.3.3.4) catalyzes the oxidation of protoporphyrinogen IX to protoporphyrin IX. After the oxidation of protoporphyrinogen IX to protoporphyrin IX, protoporphyrin IX binds with magnesium by Mg-chelatase to synthesize chlorophyll, and it binds with iron by Fe-chelatase to synthesize heme.
Therefore, when PPO activity is inhibited, synthesis of chlorophylls and heme is inhibited and the substrate protoporphyrinogen IX leaves the normal porphyrin biosynthetic pathway, resulting in the rapid export of protoporphyrinogen IX from the chloroplast to the cytoplasm, and cytoplasmic protoporphyrin IX accumulation caused by the oxidation. Protoporphyrin IX thus accumulated generates highly reactive singlet oxygen (1O2) in the presence of light and oxygen molecules to destroy cell membrane, rapidly leading to plant cell death. Based on this principle, herbicides inhibiting PPO activity have been developed. Until now, there have been 9 families of herbicides, including pyrimidinediones, diphenyl-ethers, phenylpyrazoles, N-phenylphthalimides, thiadiazoles, oxadiazoles, triazolinones, oxazolidinediones, and others herbicides, which are classified according to their chemical structures.
Further, in order to prevent effects of these herbicides on the growth of crops while using the herbicides, there is a need to provide herbicide resistance for the crops.
Meanwhile, as photosynthetic organisms, algae have the capacity to transform sunlight into energy that can be used to synthesize a variety of useful compounds. Through photosynthetic carbon fixation, algae can convert CO2 to sugar, starch, lipids, fats, or other biomolecules, for example, thereby removing a greenhouse gas from the atmosphere. Furthermore, large scale cultures of algae can be used to produce a variety of biomolecules for use as industrial enzymes, therapeutic compounds and proteins, nutritional products, commercial products, or fuel products, for example.
A major problem in the large scale cultures of algae in bioreactors or open or closed ponds is that they can become contaminated by other, highly competitive but unwanted species of algae fungi and bacteria, as well as by rotifers and other zooplankton that devour the desired species in the cultures.
Thus, there is a need to confer herbicide resistance to the algae, in order for the herbicide resistant algae to be able to grow in the presence of the herbicide at a concentration that deters growth of competing organisms not harboring the herbicide resistance gene.